Inadequate folate in pregnant women has been linked to a number of abnormalities of human embryonic and fetal development having major impacts on both the newborn and society.
One of the most well documented are neural tube defects (“NTD”) which arise from various abnormalities of tube closure during the fourth week of pregnancy. Two main examples of NTD are spina bifida (failure of posterior closure) and anencephaly (lack of a complete brain due to failure of anterior closure) among others. Most fetuses with anencephaly are not born alive, and the rest die shortly after birth. The majority of children with spina bifida survive to become adults. However, they face many challenges among which can be hydrocephalus, Chiari II malformation, tethered spinal cord, urinary tract disorders, and learning disabilities. Many may have ambulatory problems, loss of sensation, limb deformities, and loss of muscle tone. When the malformation is located higher in the spine, the risk of multiple complications is greater. It is not unusual for patients with NTD to undergo one or more operations to alleviate these issues, such as spinal surgery and/or placement of a shunt to relieve intracranial pressure to prevent brain damage due to hydrocephalus. The average total lifetime cost of caring for a child born with spina bifida is over $500,000. There are an estimated 70,000 patients in the United States with spina bifida, and about 3,000 births each year are affected by neural tube defects. According to the March of Dimes Global Report on Birth Defects (2006), worldwide, there were 324,000 births in 2001 affected by neural tube defects, with the majority of these occurring in India and China.
Women who have had one or more NTD affected pregnancies are at about ten times higher risk of a recurrence than the general population. An especially strong risk factor is low folate intake. One indication of low folate intake would be low intake of fruits, vegetables, and fortified cereals. Moreover, polymorphisms in several folate-related genes, for example, methylenetetrahydrofolate reductase (“MTHFR”), also appear to increase risk. Starting in the early 1980's, a number of clinical trials, some randomized and placebo controlled, demonstrated that periconceptual supplementation with folic acid considerably reduces the risk of NTD. An often quoted trial of secondary prevention was performed by the Medical Research Council (“MRC”) Vitamin Study Group (“Prevention of neural tube defects: results of the Medical Research Council Vitamin Study. MRC Vitamin Study Research Group,” Lancet, 338:131-137 (1991)) in which the treatment with 4 mg/day of folic acid prevented 72% of recurrent NTD compared to the control group. Not all NTD can be prevented by folic acid. Thus, the extent of the risk reduction depends, in part, on the incidence of NTD in a given population. For example, two regions in China participated in a trial of 400 μg/d of folic acid. Before intervention, the northern and southern regions had rates of 48 and 10 NTD cases per 10,000 births, respectively. After folic acid treatment these decreased to 10 and 6 per 10,000, respectively, indicating that a greater risk reduction can be obtained with populations having higher initial risk. In countries that have adopted mandatory fortification with folic acid or in clinical trials for primary prevention of neural tube defects, the rate of neural tube affected births has been decreased to a level not less than about 4 per 10,000 births or 7 per 10,000 births and abortions. These folate non-responsive cases are considered to be an outcome of multifactorial causes of NTD.
Although a number of studies have indicated that women who are folate deficient are at increased risk for NTD, it has also been shown in a population of Irish subjects that simply having average folate levels is not fully protective; a projection of NTD risk versus red cell folate concentration indicated that the optimum level was at least 906 nmol/L. Thus, it has been considered that women who have had an affected birth may require higher than normal levels to overcome one or more barriers to full utilization of folate. The above-mentioned MTHFR polymorphism, although representing only a small portion of the population at risk, is perhaps one such barrier. The majority of risk cannot yet be attributed to any single genetic defect. In mouse models, there are now over 200 genes that, when disabled, promote neural tube defects, and some of these can be overcome by administration of folate. With regard to the level of folate needed to minimize risk for NTD, it should be noted that the folate in red cells is generally not considered to be readily exchanged with tissues. Red cell folate is thought to be an indicator of the average folate levels over the last several months since it is not subject to recent and transient increases due to dietary intake. However, it is the plasma that can supply folate to various organs. In particular, shortly after implantation, the trophoblast of the human embryo integrates with the endometrium of the mother to provide a path for nutrient uptake from the plasma. This initial system will not develop into a complete placenta until well after neural tube closure. From the standpoint of the developing embryo, what is important is not necessarily the red cell folate concentration, but rather the level of plasma folate averaged over each day. In the early stages of embryonic development the degeneration of decidual cells in the endometrium shortly after implantation also can supply nutrients. Again, however, the nutrient content (such as folate) of such cells is more likely to reflect recent plasma concentrations than would red cell folate. Since a dose of folate produces a transient increase in the blood that clears over several hours, a fasting plasma sample that is remote from the last intake of folate represents a level close to the minimum for the day.
Several trials (both primary and secondary) have examined the reduction of risk for NTD at various doses of folic acid. The group treated with 400 μg/d folic acid in the trial northern and southern China reduced incidence of NTD to a level about as low as seen in other studies at higher doses. From this, many have concluded that long term supplementation with 400 μg/d is sufficient to almost fully remove the folate dependent risk for NTD. In addition, several studies have examined the folate levels achieved with long term folic acid supplementation. The plateau value of plasma folate achieved after many months of consumption of 400 μg/d is between 35 nM to more typically near 50 nM.
The current Recommended Daily Intake (“RDI”) in the U.S. for folate is 400 μg/d for the general population and 800 μg/d for pregnant and lactating women. Several public health agencies advise that all women of child bearing age consume a daily supplement containing 400 μg of folic acid regardless of specific plans for becoming pregnant. This is based on the understanding that neural tube closure is typically complete by about 28 days after conception. A study of the rate at which 400 μg/d of folic acid or 5-methyltetrahydrofolate increases folate levels when administered to young women showed that a plateau was reached in the plasma only after 12 weeks. Another study with a daily dose of 800 μg indicated that red cell concentrations could be elevated to what has been considered to be the fully protective level of 906 nM only after 4.2±3.5 weeks of treatment (Daly et al., “Folate Levels and Neural Tube Defects. Implications for Prevention,” JAMA, 274(21):1698-1702 (1995)). A report by Nguyen et al, “Steady State Folate Concentrations Achieved with 5 Compared with 1.1 Mg Folic Acid Supplementation Among Women of Childbearing Age,” Am. J. Clin. Nut., 89:844-852 (2009), showed that a group of women, who at baseline were already well above a red cell folate level of 1,000 nM, could be made even more replete by 2 weeks when given 1.1 or 5 mg/d of folic acid. The Upper Tolerable Limit for folic acid has been set to be 1.0 mg based primarily on the potential adverse interaction of high folate levels concurrent with undiagnosed vitamin B12 deficiency. Thus, it is widely believed that supplementation with folic acid should start at least several months prior to planning a pregnancy. However, only about one third of women of child bearing age in the U.S. consume supplements containing folic acid daily. Moreover, about one half of pregnancies are unplanned.
As a result, many countries have established mandatory fortification with folic acid based largely on the initial experience in the United States. Folic acid is typically added to a widely consumed grain, such as wheat, to expose the population to an average level based on the typical pattern of intake. This approach has been shown to increase the average folate levels of the population while at the same time decreasing the risk for NTD. Despite the success of these fortification programs, several groups remain unprotected. First, many countries, for example those within Europe, have not introduced mandatory fortification. Part of the debate preventing implementation is that mandatory fortification exposes the entire population to folic acid when the target is primarily only women of child bearing age. High folic acid intake by those who are vitamin B12 deficient, e.g., especially the elderly, can exacerbate neurologic damage. In several countries, even voluntary fortification, such as of breakfast cereals, is either minimal or banned. People in these areas must rely almost exclusively on the endogenous natural folate in the food they eat, and can have relatively low blood folate levels if not consuming supplements. Second, even in the countries having folic acid fortification, such as the U.S., there are subpopulations of women of child bearing age who have well below average folate levels. Such women would not be considered folate deficient according to criteria related to minimizing anemia or elevated homocysteine. However, as mentioned above, higher than average folate levels is required to fully decrease the risk of NTD affected births in those who are susceptible. According to the currently accepted thinking, once a woman has been determined to be pregnant and also to have low blood folate levels, it is considered too late to initiate administration of folic acid to meaningfully decrease the chance of having an NTD affected birth. She would, nonetheless, typically be prescribed a folic acid containing prenatal supplement to avert other problems later in pregnancy.
Inadequate folate in pregnant women has also been linked to congenital heart defects. Congenital heart defects, which affect almost 1% of live births, have a substantial impact on the viability and health of the newborn. It is one of the most common birth defects, affecting about 35,000 children a year in the United States alone, and more than 1 million world-wide. Defects of the ventricular outflow tract are a major contributor to cardiac conotruncal anomalies. It has been found that migration and integration of cells from the neural crest is essential for septation of the outflow tract and development of the aortic arch arteries. In humans migration of the cardiac neural crest cells commences as the neural tube is closing, and then enter the aortic arches and nascent outflow tracts between 32 to 37 days. By about 8 weeks after conception the outflow tract and ventricular septation is complete.
A number of studies have indicated a protective role for folate in preventing congenital heart defects, and a recent review concluded that periconceptional use of folic acid was associated with 20% decrease in the prevalence of heart defects. Mandatory fortification of flour with folic acid in Canada starting in 1998 has been observed to be related to a sharp decrease in heart defects in Quebec Province.
Similar to the heart, much of the prechordal plate that eventually gives rise to the mouth also arises from migration of neural crest cells. The lip normally closes by 35 to 40 days after conception, and the palate by 7 to 10 weeks. Orofacial clefts are common birth defects which can be classified as improper closure of the lip or palate or both. In the United States, over 4,200 and 2,600 babies with cleft lip/palate and cleft lip only are born each year, respectively. A smaller number of births are affected by orofacial clefts in conjunction with other defects (syndromic clefts).
The precise knowledge of the effect of periconceptual folic acid administration on orofacial clefts is limited by a deficiency of randomized controlled trials. However, several intervention trials have indicated a reduction in risk for recurrence, especially of cleft lip/palate. A meta-analysis of these results suggested about a two thirds reduction in recurrence. A relative risk reduction of 0.25 has been reported for non-syndromic cleft lip/palate in a study in northern China.
For each of the above heart, lip/palate, and especially neural tube birth defects, there is evidence that administration of folic acid can reduce the risk of occurrence or reoccurrence. The current and widespread understanding is that this folic acid must be given sufficiently in advance of pregnancy by many weeks if not several months to be effective.
Adequate folate during pregnancy has been linked to several other conditions.
For example, observational studies and clinical trials have indicated that folate intake during pregnancy is associated with higher birthweight. The risk of morbidity and death in infancy is increased by low birthweight. Folate levels can also influence methylation patterns, alteration of which has also recently been shown to be associated with birthweight.
A relationship has been observed between increased risk for autism and a interpregnancy interval shorter than 24 months. For example, a child born within 12 months of the previous sibling was over three times as likely to be diagnosed with autism spectrum disorder than the most spaced (>3 years) group. It was suggested that folate depletion of the mother during a given pregnancy might carry over to the next if not adequately spaced. More recently, a mother's use of prenatal vitamins prior to pregnancy or within the first month of pregnancy has been correlated with decreased risk of autism. This correlation was especially strong in women having the MTHFR TT or cystathionine β-synthase (GT or TT) genotypes. Although the hypothesis that the mother's folate levels is related to risk of autism in her offspring remains to be confirmed, the current thinking is that this may be a factor in decreasing the risk for autism.
In view of the above discussion, there is a great unmet need for a way to decrease the occurrence of birth defects among women who do not have optimal folate levels and to reduce the risk of NTD and other birth defects and disorders/pathologies among women who are found to be pregnant while possibly having inadequate plasma folate and/or other risk factors. The present invention, at least in part, is meant to address these needs.